Clostridium difficile (hereinafter may be referred to as “C. difficile”) is a sporulating Gram-positive bacillus which produces an exotoxin pathogenic to human. In recent years, C. difficile-associated diarrhea (CDAD) caused by this bacterium has become a serious problem (Non-Patent Document 1). Specifically, overuse of antibiotics, anticancer agents, etc. causes damage to normal intestinal flora, and the thus-grown C. difficile produces toxins TcdA and TcdB, to thereby develop symptoms such as diarrhea. As has been known, C. difficile is excreted in feces from the infected individual, and other individuals are infected with C. difficile through the mouth or mucosa thereof via, for example, instruments or hands.
The pathogenicity of C. difficile is generally attributed to two toxins TcdA and TcdB belonging to the large clostridial toxins (LCTs) family. Strains of C. difficile are roughly classified, according to the difference in toxigenicity therebetween, into a type of TcdA production and TcdB production (A+B+), a type of no TcdA production and TcdB production (A−B+), and a type of no toxin production (A−B−). The toxin production system of C. difficile is considered to include tcdA and tcdB, and a pathogenic locus formed by tcdC, tcdR, and tcdE which encode the regulatory factors of tcdA and tcdB. As has been reported, when tcdC which negatively controls toxin production is deleted, production of the toxins TcdA and TcdB is enhanced.
Therefore, selective detection of only toxigenic C. difficile strains (A+B+ and A−B+) is clinically important for diagnosis of a Clostridium difficile infection (CDI) such as CDAD.
Hitherto, it has been reported that several primers targeting the toxin genes tcdA and tcdB are employed for detection of toxigenic C. difficile, and the genes can be detected in DNA extracted from feces by means of these primers (Non-Patent Documents 2 to 4 and Patent Document 1).
However, since the number of C. difficile cells is small in the healthy human bowel, detection of toxigenic strains in feces by PCR requires establishment of a detection system having high specificity and detection sensitivity. Thus, conventional techniques are not satisfactory from this viewpoint.